Decitabine suppresses triple-negative breast cancer by disrupting DNMT1 liquid-liquid phase separation and synergizes with G9a inhibition.

Triple-negative breast cancer (TNBC) remains a substantial clinical challenge due to the lack of well-defined therapeutic targets. Previous studies have demonstrated that epigenetic modifiers play crucial roles in the progression of various cancers. In this study, we identify DNA methyltransferase 1 (DNMT1) as a critical epigenetic driver of TNBC proliferation and metastasis. Our findings show that DNMT1 is highly expressed in TNBC and forms liquid-liquid phase separation (LLPS) condensates. Knockout of DNMT1 sensitizes TNBC cells to apoptosis and lipid peroxidation, downregulates oncogenic pathways, and upregulates apoptosis-related genes. The DNMT1 inhibitor decitabine (DAC) disrupts DNMT1 condensates, consequently inducing apoptosis and lipid peroxidation. Mechanistically, DAC directly binds to the disordered domain of DNMT1, thereby dissolving LLPS condensates and impairing epigenetic silencing. Remarkably, DAC synergizes with histone methyltransferase G9a inhibitor BIX-01294 (BIX) to suppress TNBC proliferation, migration, and invasion. Combined treatment with DAC and BIX significantly inhibits lung metastasis in murine and human TNBC mouse models, reducing proliferative cancer cells and metastatic burden. This study reveals DNMT1 LLPS as a druggable vulnerability in TNBC and establishes LLPS disruption as a promising therapeutic strategy, providing a rationale for combining epigenetic inhibitors to treat TNBC and other cancers with high DNMT1 expression.